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Optogenetically activated astrocytes induced excitatory response in SF-1 neurons, mainly via NMDA receptor. (a) Schematic representing viral construct of ChR2 specifically expressed in astrocytes. (b) Whole-cell recording of VMH neurons near ChR2-expressing astrocytes. (c) Representative recordings illustrating VMH SF-1 neurons depolarized by optical stimulation of astrocytes. (d) Application of glutamatergic receptor antagonists <t>NBQX</t> (50 μ M) <t>and</t> <t>D-APV</t> (50 μ M) suppressed action potential firing caused by optical stimulation of astrocytes, which was recovered after aCSF washout ( n = 6). (e) Application of D-APV significantly diminished membrane potential depolarization induced by astrocyte activation ( n = 5, ∗∗ p < 0.01; paired t -test). (f) NBQX did not influence interaction between astrocytes and SF-1 neurons ( n = 5, ∗ p < 0.05; paired t -test). (g) Schematic showing that manipulation of astrocytes in VMH could bidirectionally regulate chronic stress-induced anxiety and bone loss by affecting neural excitatory through NMDA receptors.
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Optogenetically activated astrocytes induced excitatory response in SF-1 neurons, mainly via NMDA receptor. (a) Schematic representing viral construct of ChR2 specifically expressed in astrocytes. (b) Whole-cell recording of VMH neurons near ChR2-expressing astrocytes. (c) Representative recordings illustrating VMH SF-1 neurons depolarized by optical stimulation of astrocytes. (d) Application of glutamatergic receptor antagonists <t>NBQX</t> (50 μ M) <t>and</t> <t>D-APV</t> (50 μ M) suppressed action potential firing caused by optical stimulation of astrocytes, which was recovered after aCSF washout ( n = 6). (e) Application of D-APV significantly diminished membrane potential depolarization induced by astrocyte activation ( n = 5, ∗∗ p < 0.01; paired t -test). (f) NBQX did not influence interaction between astrocytes and SF-1 neurons ( n = 5, ∗ p < 0.05; paired t -test). (g) Schematic showing that manipulation of astrocytes in VMH could bidirectionally regulate chronic stress-induced anxiety and bone loss by affecting neural excitatory through NMDA receptors.
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(A) Typical electrode placement for synaptic stimulation. The picture shows an infra-red differential interference contrast image of a horizontal slice from the hippocampus, where the stimulating electrode (s.e.) is placed in the molecular layer (m.l.) and an extracellular recording electrode (r.e.) is inserted in the layer of dentate granule neurons (DGNs, following the arrow direction). (B) Inhibition of ionotropic glutamate receptors abolishes the metabolic transients. A representative trace of a synaptically stimulated DGN (using a train of 60 pulses at 20 Hz) shows that the Peredox signal disappears after blocking synaptic transmission with a mixture of 5 μM <t>NBQX</t> and 25 <t>μM</t> <t>D-AP5</t> in the ACSF. The complete blockade of synaptic transmission is corroborated by the absence of the typical RCaMP1h signal after stimulation. The effect of synaptic blockers was partially reversed after 5 min of washout in regular ACSF, and both Ca2+ and NADH transients were completely recovered after 10 min in ACSF. (C) Dataset for the effect of ionotropic glutamate receptor block. Data were compared by a paired Student’s t-test (neurons=46, slices=6, mice=3). (D) Typical electrode placement for antidromic (axonal) stimulation. In this experimental paradigm, the stimulating electrode (s.e.) is placed in the the hilus (hil) and an extracellular recording electrode (r.e.) is inserted in the layer of dentate granule neurons (DGNs, following the arrow direction). (E) Average transient increases in Peredox lifetime after antidromic stimulation of 100 brief pulses delivered at 50 Hz. Traces represent the mean ± SEM (events=179, neurons=84, slices=18, mice=11; SEM calculated using number of neurons). The lower panel shows the putative average Ca2+ spike in the cells, as a proxy for neuronal excitation. (F) Elevation in the NADH/NAD+ ratio is directly correlated with the neuronal Ca2+ increase after electrical antidromic stimulation in DGNs, similarly to synaptic stimulation. Scatter plots of changes in Peredox lifetime versus variations in RCaMP lifetime for events recorded after antidromic stimulation (open circles; events=205, neurons=87, slices=19, mice=12), overlap with data collected using synaptic stimulation (dark squares).
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(A) Typical electrode placement for synaptic stimulation. The picture shows an infra-red differential interference contrast image of a horizontal slice from the hippocampus, where the stimulating electrode (s.e.) is placed in the molecular layer (m.l.) and an extracellular recording electrode (r.e.) is inserted in the layer of dentate granule neurons (DGNs, following the arrow direction). (B) Inhibition of ionotropic glutamate receptors abolishes the metabolic transients. A representative trace of a synaptically stimulated DGN (using a train of 60 pulses at 20 Hz) shows that the Peredox signal disappears after blocking synaptic transmission with a mixture of 5 μM <t>NBQX</t> and 25 <t>μM</t> <t>D-AP5</t> in the ACSF. The complete blockade of synaptic transmission is corroborated by the absence of the typical RCaMP1h signal after stimulation. The effect of synaptic blockers was partially reversed after 5 min of washout in regular ACSF, and both Ca2+ and NADH transients were completely recovered after 10 min in ACSF. (C) Dataset for the effect of ionotropic glutamate receptor block. Data were compared by a paired Student’s t-test (neurons=46, slices=6, mice=3). (D) Typical electrode placement for antidromic (axonal) stimulation. In this experimental paradigm, the stimulating electrode (s.e.) is placed in the the hilus (hil) and an extracellular recording electrode (r.e.) is inserted in the layer of dentate granule neurons (DGNs, following the arrow direction). (E) Average transient increases in Peredox lifetime after antidromic stimulation of 100 brief pulses delivered at 50 Hz. Traces represent the mean ± SEM (events=179, neurons=84, slices=18, mice=11; SEM calculated using number of neurons). The lower panel shows the putative average Ca2+ spike in the cells, as a proxy for neuronal excitation. (F) Elevation in the NADH/NAD+ ratio is directly correlated with the neuronal Ca2+ increase after electrical antidromic stimulation in DGNs, similarly to synaptic stimulation. Scatter plots of changes in Peredox lifetime versus variations in RCaMP lifetime for events recorded after antidromic stimulation (open circles; events=205, neurons=87, slices=19, mice=12), overlap with data collected using synaptic stimulation (dark squares).
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Image Search Results


Optogenetically activated astrocytes induced excitatory response in SF-1 neurons, mainly via NMDA receptor. (a) Schematic representing viral construct of ChR2 specifically expressed in astrocytes. (b) Whole-cell recording of VMH neurons near ChR2-expressing astrocytes. (c) Representative recordings illustrating VMH SF-1 neurons depolarized by optical stimulation of astrocytes. (d) Application of glutamatergic receptor antagonists NBQX (50 μ M) and D-APV (50 μ M) suppressed action potential firing caused by optical stimulation of astrocytes, which was recovered after aCSF washout ( n = 6). (e) Application of D-APV significantly diminished membrane potential depolarization induced by astrocyte activation ( n = 5, ∗∗ p < 0.01; paired t -test). (f) NBQX did not influence interaction between astrocytes and SF-1 neurons ( n = 5, ∗ p < 0.05; paired t -test). (g) Schematic showing that manipulation of astrocytes in VMH could bidirectionally regulate chronic stress-induced anxiety and bone loss by affecting neural excitatory through NMDA receptors.

Journal: Neural Plasticity

Article Title: Astrocytes in the Ventromedial Hypothalamus Involve Chronic Stress-Induced Anxiety and Bone Loss in Mice

doi: 10.1155/2021/7806370

Figure Lengend Snippet: Optogenetically activated astrocytes induced excitatory response in SF-1 neurons, mainly via NMDA receptor. (a) Schematic representing viral construct of ChR2 specifically expressed in astrocytes. (b) Whole-cell recording of VMH neurons near ChR2-expressing astrocytes. (c) Representative recordings illustrating VMH SF-1 neurons depolarized by optical stimulation of astrocytes. (d) Application of glutamatergic receptor antagonists NBQX (50 μ M) and D-APV (50 μ M) suppressed action potential firing caused by optical stimulation of astrocytes, which was recovered after aCSF washout ( n = 6). (e) Application of D-APV significantly diminished membrane potential depolarization induced by astrocyte activation ( n = 5, ∗∗ p < 0.01; paired t -test). (f) NBQX did not influence interaction between astrocytes and SF-1 neurons ( n = 5, ∗ p < 0.05; paired t -test). (g) Schematic showing that manipulation of astrocytes in VMH could bidirectionally regulate chronic stress-induced anxiety and bone loss by affecting neural excitatory through NMDA receptors.

Article Snippet: NBQX (50 μ M, MCE, Shanghai, China) and D-APV (50 μ M, MCE Shanghai, China) were used in perfused aCSF to block the function of AMPA and NMDA receptors, respectively.

Techniques: Construct, Expressing, Membrane, Activation Assay

(A) Typical electrode placement for synaptic stimulation. The picture shows an infra-red differential interference contrast image of a horizontal slice from the hippocampus, where the stimulating electrode (s.e.) is placed in the molecular layer (m.l.) and an extracellular recording electrode (r.e.) is inserted in the layer of dentate granule neurons (DGNs, following the arrow direction). (B) Inhibition of ionotropic glutamate receptors abolishes the metabolic transients. A representative trace of a synaptically stimulated DGN (using a train of 60 pulses at 20 Hz) shows that the Peredox signal disappears after blocking synaptic transmission with a mixture of 5 μM NBQX and 25 μM D-AP5 in the ACSF. The complete blockade of synaptic transmission is corroborated by the absence of the typical RCaMP1h signal after stimulation. The effect of synaptic blockers was partially reversed after 5 min of washout in regular ACSF, and both Ca2+ and NADH transients were completely recovered after 10 min in ACSF. (C) Dataset for the effect of ionotropic glutamate receptor block. Data were compared by a paired Student’s t-test (neurons=46, slices=6, mice=3). (D) Typical electrode placement for antidromic (axonal) stimulation. In this experimental paradigm, the stimulating electrode (s.e.) is placed in the the hilus (hil) and an extracellular recording electrode (r.e.) is inserted in the layer of dentate granule neurons (DGNs, following the arrow direction). (E) Average transient increases in Peredox lifetime after antidromic stimulation of 100 brief pulses delivered at 50 Hz. Traces represent the mean ± SEM (events=179, neurons=84, slices=18, mice=11; SEM calculated using number of neurons). The lower panel shows the putative average Ca2+ spike in the cells, as a proxy for neuronal excitation. (F) Elevation in the NADH/NAD+ ratio is directly correlated with the neuronal Ca2+ increase after electrical antidromic stimulation in DGNs, similarly to synaptic stimulation. Scatter plots of changes in Peredox lifetime versus variations in RCaMP lifetime for events recorded after antidromic stimulation (open circles; events=205, neurons=87, slices=19, mice=12), overlap with data collected using synaptic stimulation (dark squares).

Journal: Cell metabolism

Article Title: Neuronal stimulation triggers neuronal glycolysis and not lactate uptake

doi: 10.1016/j.cmet.2017.06.021

Figure Lengend Snippet: (A) Typical electrode placement for synaptic stimulation. The picture shows an infra-red differential interference contrast image of a horizontal slice from the hippocampus, where the stimulating electrode (s.e.) is placed in the molecular layer (m.l.) and an extracellular recording electrode (r.e.) is inserted in the layer of dentate granule neurons (DGNs, following the arrow direction). (B) Inhibition of ionotropic glutamate receptors abolishes the metabolic transients. A representative trace of a synaptically stimulated DGN (using a train of 60 pulses at 20 Hz) shows that the Peredox signal disappears after blocking synaptic transmission with a mixture of 5 μM NBQX and 25 μM D-AP5 in the ACSF. The complete blockade of synaptic transmission is corroborated by the absence of the typical RCaMP1h signal after stimulation. The effect of synaptic blockers was partially reversed after 5 min of washout in regular ACSF, and both Ca2+ and NADH transients were completely recovered after 10 min in ACSF. (C) Dataset for the effect of ionotropic glutamate receptor block. Data were compared by a paired Student’s t-test (neurons=46, slices=6, mice=3). (D) Typical electrode placement for antidromic (axonal) stimulation. In this experimental paradigm, the stimulating electrode (s.e.) is placed in the the hilus (hil) and an extracellular recording electrode (r.e.) is inserted in the layer of dentate granule neurons (DGNs, following the arrow direction). (E) Average transient increases in Peredox lifetime after antidromic stimulation of 100 brief pulses delivered at 50 Hz. Traces represent the mean ± SEM (events=179, neurons=84, slices=18, mice=11; SEM calculated using number of neurons). The lower panel shows the putative average Ca2+ spike in the cells, as a proxy for neuronal excitation. (F) Elevation in the NADH/NAD+ ratio is directly correlated with the neuronal Ca2+ increase after electrical antidromic stimulation in DGNs, similarly to synaptic stimulation. Scatter plots of changes in Peredox lifetime versus variations in RCaMP lifetime for events recorded after antidromic stimulation (open circles; events=205, neurons=87, slices=19, mice=12), overlap with data collected using synaptic stimulation (dark squares).

Article Snippet: KEY RESOURCES TABLE REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Bacterial and Virus Strains AAV9.Syn.RCaMP1h.WPRE.SV40 Penn Vector Core Discontinued AAV9.Syn.jRCaMP1b-NES.WPRE.SV40 Penn Vector Core Cat#AV-9-PV3852 Biological Samples Chemicals, Peptides, and Recombinant Proteins NBQX (6-Nitro-7-sulfamoylbenzo[f]quinoxaline-2,3-dione, Disodium Salt) Toronto Research Chemicals Cat#N550005; CAS:479347-86-9 D-AP5 (D-(-)-2-Amino-5-phosphonopentanoic acid) Abcam Cat#ab120003; CAS:79055-68-8 AR-C155858 Tocris Bioscience Cat#4960; CAS:496791-37-8 Picrotoxin Sigma-Aldrich Cat#P1675; CAS:124-87-8 Cytochalasin B Tocris Bioscience Cat#5474; CAS:14930-96-2 Cytochalasin D Tocris Bioscience Cat#1233; CAS:22144-77-0 GSK-2837808A Tocris Bioscience Cat#5189; CAS:1445879-21-9 Aminooxyacetate (O-(carboxymethyl)hydroxylamine hemihydrate) Sigma-Aldrich Cat#C13408; CAS:2921-14-4 Sodium L-lactate Sigma-Aldrich Cat#71718; CAS:867-56-1 Sodium pyruvate Sigma-Aldrich Cat#P8574; CAS:113-24-6 Critical Commercial Assays Deposited Data Experimental Models: Cell Lines Experimental Models: Organisms/Strains C57BL/6NCrl mice Charles River RRID:IMSR_CRL:27 Oligonucleotides Recombinant DNA AAV.CAG.Peredox.WPRE.SV40 Mongeon et al., 2016 Addgene #73807 AAV.hSyn.Peredox-NLS.WPRE.SV40 This paper Derived from Addgene #32384 AAV.CAG.SweetieTS.WPRE.SV40 This paper AAV.CAG.Laconic.WPRE.SV40 San Martín et al., 2013 Derived from Addgene #44238 AAV.CAG.pHRed.WPRE.SV40 Tantama et al., 2011 Derived from Addgene #31472 Software and Algorithms GraphPad Prism Version 7 GraphPad Origin 2015 OriginLab ScanImage Pologruto et al., 2003 Other Polymeric microporous filters with 8μm pores and 25 μm thick grid structural support Precision Membranes, Provo, UT N/A Open in a separate window Quantification and Statistical Analyses

Techniques: Inhibition, Blocking Assay, Transmission Assay

Quantification and Statistical Analyses

Journal: Cell metabolism

Article Title: Neuronal stimulation triggers neuronal glycolysis and not lactate uptake

doi: 10.1016/j.cmet.2017.06.021

Figure Lengend Snippet: Quantification and Statistical Analyses

Article Snippet: KEY RESOURCES TABLE REAGENT or RESOURCE SOURCE IDENTIFIER Antibodies Bacterial and Virus Strains AAV9.Syn.RCaMP1h.WPRE.SV40 Penn Vector Core Discontinued AAV9.Syn.jRCaMP1b-NES.WPRE.SV40 Penn Vector Core Cat#AV-9-PV3852 Biological Samples Chemicals, Peptides, and Recombinant Proteins NBQX (6-Nitro-7-sulfamoylbenzo[f]quinoxaline-2,3-dione, Disodium Salt) Toronto Research Chemicals Cat#N550005; CAS:479347-86-9 D-AP5 (D-(-)-2-Amino-5-phosphonopentanoic acid) Abcam Cat#ab120003; CAS:79055-68-8 AR-C155858 Tocris Bioscience Cat#4960; CAS:496791-37-8 Picrotoxin Sigma-Aldrich Cat#P1675; CAS:124-87-8 Cytochalasin B Tocris Bioscience Cat#5474; CAS:14930-96-2 Cytochalasin D Tocris Bioscience Cat#1233; CAS:22144-77-0 GSK-2837808A Tocris Bioscience Cat#5189; CAS:1445879-21-9 Aminooxyacetate (O-(carboxymethyl)hydroxylamine hemihydrate) Sigma-Aldrich Cat#C13408; CAS:2921-14-4 Sodium L-lactate Sigma-Aldrich Cat#71718; CAS:867-56-1 Sodium pyruvate Sigma-Aldrich Cat#P8574; CAS:113-24-6 Critical Commercial Assays Deposited Data Experimental Models: Cell Lines Experimental Models: Organisms/Strains C57BL/6NCrl mice Charles River RRID:IMSR_CRL:27 Oligonucleotides Recombinant DNA AAV.CAG.Peredox.WPRE.SV40 Mongeon et al., 2016 Addgene #73807 AAV.hSyn.Peredox-NLS.WPRE.SV40 This paper Derived from Addgene #32384 AAV.CAG.SweetieTS.WPRE.SV40 This paper AAV.CAG.Laconic.WPRE.SV40 San Martín et al., 2013 Derived from Addgene #44238 AAV.CAG.pHRed.WPRE.SV40 Tantama et al., 2011 Derived from Addgene #31472 Software and Algorithms GraphPad Prism Version 7 GraphPad Origin 2015 OriginLab ScanImage Pologruto et al., 2003 Other Polymeric microporous filters with 8μm pores and 25 μm thick grid structural support Precision Membranes, Provo, UT N/A Open in a separate window Quantification and Statistical Analyses

Techniques: Plasmid Preparation, Recombinant, Derivative Assay, Software

Journal: Neuron

Article Title: Activity-Dependent Plasticity of Axo-axonic Synapses at the Axon Initial Segment

doi: 10.1016/j.neuron.2020.01.037

Figure Lengend Snippet:

Article Snippet: NBQX , Santa Cruz Biotechnology , sc-222048; CAS: 479347-86-9.

Techniques: Virus, Plasmid Preparation, Recombinant, Software, Fluorescence, Imaging